Mantle cell lymphoma (MCL) remains incurable due to the development of drug resistance despite advances in targeted therapy. Each successive treatment failure is associated with a more rapidly proliferating disease and fewer practical treatment options. For example, the BTK inhibitor (BTKi) ibrutinib has unprecedented activity but failure is virtually universal and is associated with dismal outcomes. Understanding the mechanism and genomic basis for therapy resistance is thus fundamental to developing superior durable therapies that are also amenable to patient stratification in MCL. As dysregulation of CDK4 (CDK6 is not expressed in MCL) and aberrant cyclin D1 expression underlie MCL proliferation, targeting CDK4 represents a rational approach to developing novel therapies that circumvent ibrutinib resistance in MCL. We have previously demonstrated that the clinical response to ibrutinib was tightly associated with PI3K inactivation in MCL patients. Induction of prolonged early G1 arrest (pG1) by inhibiting CDK4 with palbociclib (selective CDK4/6 inhibitor) restricts the expression of genes to those scheduled for early G1 only. This led to an imbalance in gene expression that reprogrammed MCL cells for therapeutic vulnerability, including BTK and PI3K inhibition. On this basis, we hypothesize that targeting CDK4 will both restrict the expansion of resistant clones and reprogram MCL cells for a deeper, and more durable clinical response to BTKi or PI3Ki. Supporting this hypothesis, our phase I clinical trial of palbociclib in combination with ibrutinib for patients with recurrent MCL (PALIBR) revealed a promising 67% overall response rate with 43% complete response (N=27), Moreover the response was rapid and durable; only 14% responding patients have progressed in ~32 months since the trial opened. Capitalizing on these exciting findings and the upcoming phase 2 PALIBR clinical trial, we propose to define the mechanism of pG1 reprograming for therapeutic vulnerability and the molecular biomarkers that discriminate sensitivity from resistance to targeting CDK4 in MCL with two specific aims: 1) to determine the genomic basis for resistance to targeting CDK4 in combination therapy. We will determine if composite copy number variation (CNV) and clonal selection causes resistance to PALIBR by longitudinal functional genomics and develop strategies to circumvent PALIBR resistance by targeting the therapeutic vulnerability conferred by resistance biomarkers; 2) to elucidate the mechanism for pG1 reprogramming in MCL cells. We will test our hypothesis that by reinforcing Rb sequestration of E2F1, CDK4 inhibition disrupts the E2F1- EZH2 regulatory circuitry, which sustains the pG1 state needed for a durable clinical response and reprograms MCL cells for therapy vulnerability through chromatin remodeling, repression of IRF4 and inactivation of PI3K. Targeting CDK4/6 is now FDA-approved for treatment of breast cancer and is being actively investigated in diverse human cancers, yet the underlying mechanism remains unknown. Successful completion of the proposed studies should both advance MCL therapy and have a profound impact on targeting the cell cycle in human cancer.